ELECTRON-PARAMAGNETIC-RESONANCE STUDIES OF THE REACTION OF ARYL RADICALS WITH NUCLEIC-ACIDS AND THEIR COMPONENTS

Aryl radicals may be responsible for the DNA damage observed in both c ellular systems and isolated DNA exposed to a number of systems (such as benzoyl peroxide or arenediazonium ion/metal ion couples) which are believed to be capable of generating such species, though it is uncle ar how this damage arises. EPR spectroscopy in conjunction with spin t rapping [using 2-methyl-2-nitrosopropane (MNP)I has therefore been uti lised to study the mechanism and sites of attack of aryl radicals (gen erated by treatment of the corresponding diazonium ions with Fe2+-EDTA or Ti3+) on DNA, RNA and their components. The results obtained sugge st that, for the pyrimidine nucleobases, nucleosides and nucleotides, the major mode of reaction is addition to the alkenic C-5-C-6 double b ond of the base moiety, though significant yields of other radicals, b elieved to arise from abstraction of hydrogen at the sugar moiety, are also observed with some of the nucleosides and nucleotides. Radicals arising from attack on adenosine 5'-triphosphate have also been detect ed. The increased yield of sugar-derived radicals in these reactions, when compared with those previously reported for (electrophilic) HO. a nd alkoxyl radicals, is in accord with the known nucleophilic nature o f aryl radicals. Studies with the polyU, polyA . polyU, polyC, RNA and DNA suggest that aryl radicals also damage these macromolecules, thou gh the broad nature of the spectral lines and interference from the si gnal of the aryl-radical adduct to the spin trap prevent detailed iden tification of the site(s) of attack. For DNA and RNA the signals obtai ned are pH dependent. At pH 7.4 both slowly tumbling and rapidly tumbl ing spin adducts are observed with tRNA, which is consistent with the spin trapping of both large, substrate-derived, radicals and low-molec ular-mass fragments, possibly from the sugar moieties. With DNA only s pectra from rapidly tumbling species are seen at pH 7.4; these are aga in believed to be due to the presence of low-molecular-mass material. The formation of these small fragments suggests that aryl radicals are capable of generating strand breaks in nucleic acids, and therefore t hat such species may be responsible for the genetic damage observed in cells exposed to aryl-radical-generating systems.